The NBDs that wouldn't die: A cautionary tale of the use of isolated nucleotide binding domains of ABC transporters

A - Papers appearing in refereed journals

De Marcos Lousa, C., Dietrich, D., Johnson, B., Baldwin, S. A., Holdsworth, M. J., Theodoulou, F. L. and Baker, A. 2009. The NBDs that wouldn't die: A cautionary tale of the use of isolated nucleotide binding domains of ABC transporters. Communicative and Integrative Biology. 2 (2), pp. 97-99. https://doi.org/10.4161/cib.7621

AuthorsDe Marcos Lousa, C., Dietrich, D., Johnson, B., Baldwin, S. A., Holdsworth, M. J., Theodoulou, F. L. and Baker, A.
Abstract

COMATOSE (CTS), the plant homologue of Adrenoleukodystrophy protein, is a full length ABC transporter localised in peroxisomes. In a recent article, we reported that the two nucleotide binding domains of CTS are not functionally equivalent in vivo. Mutations in conserved residues in the Walker A (K487A) and B (D606N) motifs of NBD1 resulted in a null phenotype, whereas identical mutations in the equivalent residues in NBD2 (K1136A and D1276N) had no detectable effect.1 In order to study the effect of these mutations on the ATPase activity of the nucleotide binding domains, we cloned and expressed the isolated NBDs as maltose binding protein (MBP) fusion proteins. We show that ATPase activity is associated with the isolated MBP-NBDs. However, mutations of amino acids located in conserved motifs did not result in striking reduction in activity despite well characterized roles in ATP binding and hydrolysis. We urge caution in the interpretation of results obtained from the study of isolated NBD fusions and their extrapolation to the mechanism of ATP hydrolysis in ABC transporter proteins.

Year of Publication2009
JournalCommunicative and Integrative Biology
Journal citation2 (2), pp. 97-99
Digital Object Identifier (DOI)https://doi.org/10.4161/cib.7621
Open accessPublished as ‘gold’ (paid) open access
FunderBiotechnology and Biological Sciences Research Council
Funder project or codeCentre for Crop Genetic Improvement (CGI)
Purification and functional characterisation of COMATOSE a peroxisomal ABC transporter from Arabidopsis thaliana
Publisher's version
Copyright license
CC BY
Output statusPublished
Publication dates
Online30 Apr 2009
Publication process dates
Accepted15 Dec 2008

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